Knowledge of the significance of the diverse levels and patterns of organization of DNA sequence variation in natural populations is central to understanding genome evolution. Our proposed experiments are motivated by our finding of strongly contrasting levels of DNA restriction site and protein polymorphism between Dorscophila melanogaster and D. simulans, significant linkage disequilibrium over as much as 10 map units on chromosome 3 of D. melanogaster and a correlation across chromosomes and species between strong linkage disequilibrium and polymorphic inversions, and by the development of a statistical test of neutral molecular evolution based on the comparison on intra-specific DNA sequence variation with inter-specific divergence. We will obtain accurate estimates of intra-and interspecific DNA sequence variation in D. melanogaster and D. simulans to examine hypotheses to explain the patterns of variation observed within and between these species. This includes a 4-cutter restriction site analysis of the rosy, 1(3)S12 and snake genes in populations samples of these species, and the direct sequencing of a random sample of alleles from each species for the 1(3)S12 and rosy genes. We will also define the levels and physical limits of linkage disequilibrium among genetically mapped sequence variants on the third and second chromosomes of D. melanogaster and D. simulans to examine the relationship between linkage disequilibrium and region-specific variation in rates of recombination (including that due to the presence of polymorphic inversions). This will involve scoring polymorphic DNA variants using 4-cutters and PCR amplification sequencing at intervals for which empirically determined rates of recombination are available, examining patterns of linkage of disequilibrium and sequence variability within and between inversion chromosomes, and the determination of rates of recombination in inversion heterozygotes to see if they are compatible with the linkage disequilibrium observed or whether some form of selection need be invoked.